公开(公告)号：US06808907B2

公开(公告)日：2004-10-26

申请号：US10105332

申请日：2002-03-26

Applicant: Tsutomu Honma ,  Etsuko Sugawa ,  Tetsuya Yano ,  Tsuyoshi Nomoto ,  Takeshi Imamura ,  Tomohiro Suzuki ,  Takashi Kenmoku

Inventor： Tsutomu Honma ,  Etsuko Sugawa ,  Tetsuya Yano ,  Tsuyoshi Nomoto ,  Takeshi Imamura ,  Tomohiro Suzuki ,  Takashi Kenmoku

IPC: C12P762

CPC classification number: C12P7/625 ,  C12M47/06

Abstract: The present invention provides a method for producing highly purified polyhydroxyalkanoate (PHA) with high yield, by removing cell components other than PHA from cells containing PHA. Moreover, the present invention provides a method for simply removing or reducing chlorine remained in the collected PHA particles, when a treatment with an oxidizing agent containing hypochlorite is performed in the production of PHA as described above. One of the above-described methods comprises a step of treating cells containing polyhydroxyalkanoate with an oxidizing agent containing at least hypochlorite, a step of separating the treated cells into a water-soluble fraction and a water-insoluble fraction, and a step of reducing chlorine remained in the water-insoluble fraction. The step of reducing chlorine may be a step of washing the water-insoluble fraction with a hot water, thiosulfate solution or polar solvent solution containing at least an organic polar solvent in which polyhydroxyalkanoate is insoluble.

Abstract translation: 本发明提供了通过从含有PHA的细胞中除去PHA以外的细胞成分而以高产率生产高纯度聚羟基链烷酸酯（PHA）的方法。 此外，本发明提供了如上所述在生产PHA中进行含有次氯酸盐的氧化剂的处理时，简单地除去或还原所收集的PHA颗粒中残留的氯的方法。 上述方法之一包括用至少含有次氯酸盐的氧化剂处理含有聚羟基链烷酸酯的细胞的步骤，将经处理的细胞分离成水溶性级分和水不溶性级分的步骤以及还原氯 保留在水不溶性部分中。 还原氯的步骤可以是用至少含有聚羟基链烷酸酯不溶的有机极性溶剂的热水，硫代硫酸盐溶液或极性溶剂溶液洗涤水不溶性部分的步骤。

公开(公告)号：US06692945B2

公开(公告)日：2004-02-17

申请号：US09754112

申请日：2001-01-05

Applicant: Gyanendra Tripathi ,  Lata Hanamantrao Mahishi ,  Turaga Venkata Naga Ramachander ,  Shuban Kishen Rawal

Inventor： Gyanendra Tripathi ,  Lata Hanamantrao Mahishi ,  Turaga Venkata Naga Ramachander ,  Shuban Kishen Rawal

IPC: C12P762

CPC classification number: C12N15/52 ,  C12N15/70 ,  C12N15/76 ,  C12P7/625

Abstract: The present invention relates to a process for the production of polyhydroxyoctanoate, said method involving construction of a multifunctional Escherichia coli—Streptomyces conjugative shuttle vector, development of a recombinant vector designated as pCAB218, which is used to transform Streptomyces lividans TK64, such that it is capable of producing polyhydroxyoctanoate (PHO) in substantial amounts when grown in a conventional mineral medium.

Abstract translation: 本发明涉及一种生产聚羟基辛酸酯的方法，所述方法涉及构建多功能大肠杆菌 - 链霉菌共轭穿梭载体，开发用于转化变形链霉菌TK64的重组载体，命名为pCAB218，使得它是 能够在常规矿物介质中生长时能够大量生产聚羟基辛酸酯（PHO）。

公开(公告)号：US06593116B1

公开(公告)日：2003-07-15

申请号：US09375975

申请日：1999-08-17

Applicant: Gjalt W. Huisman ,  Oliver P. Peoples ,  Frank A. Skraly

Inventor： Gjalt W. Huisman ,  Oliver P. Peoples ,  Frank A. Skraly

IPC: C12P762

CPC classification number: C12N9/1029 ,  C12N9/00 ,  C12N9/0004 ,  C12N9/0006 ,  C12N9/1025 ,  C12N9/13 ,  C12N9/88 ,  C12N15/90 ,  C12P7/625 ,  Y10S435/829 ,  Y10S435/831 ,  Y10S435/877

Abstract: Transgenic microbial strains are provided which contain the genes required for PHA formation integrated on the chromosome. The strains are advantageous in PHA production processes, because (1) no plasmids need to be maintained, generally obviating the required use of antibiotics or other stabilizing pressures, and (2) no plasmid loss occurs, thereby stabilizing the number of gene copies per cell throughout the fermentation process, resulting in homogeneous PHA product formation throughout the production process. Genes are integrated using standard techniques, preferably transposon mutagenesis. In a preferred embodiment wherein mutiple genes are incorporated, these are incorporated as an operon. Sequences are used to stabilize mRNA, to induce expression as a function of culture conditions (such as phosphate concentration), temperature, and stress, and to aid in selection, through the incorporation of selection markers such as markers conferring antibiotic resistance.

公开(公告)号：US06444452B1

公开(公告)日：2002-09-03

申请号：US09723711

申请日：2000-11-28

Applicant: Vilmos Keri ,  Lajos Deak ,  Ilona Forgacs ,  Csaba Szabo ,  Edit Arvai Nagyne

Inventor： Vilmos Keri ,  Lajos Deak ,  Ilona Forgacs ,  Csaba Szabo ,  Edit Arvai Nagyne

IPC: C12P762

CPC classification number: C12P17/06 ,  C07C67/52 ,  C07C69/013 ,  C07C69/33 ,  C07C2602/28 ,  C12P7/42

Abstract: A novel process for recovering a compound from a fermentation broth that includes the stages of forming an enriched solution of the compound by extraction, obtaining a salt of the compound from the enriched solution, purifying a salt of the compound and trans-salifying the salt of the compound to a metal salt of the compound is disclosed.

Abstract translation: 一种用于从发酵液中回收化合物的新方法，其包括通过萃取形成化合物的富集溶液的阶段，从富集溶液中获得化合物的盐，纯化化合物的盐和将盐化合物转化成盐 公开了化合物与化合物的金属盐。

公开(公告)号：US06355455B1

公开(公告)日：2002-03-12

申请号：US09462427

申请日：2000-01-10

Applicant: Gilles Pauly ,  Jean-Marc Engasser ,  Mohammed Ghoul

Inventor： Gilles Pauly ,  Jean-Marc Engasser ,  Mohammed Ghoul

IPC: C12P762

CPC classification number: C12P7/64 ,  C12P19/02 ,  C12P19/44

Abstract: A method for the enzymatic synthesis of sucrose ester, comprises introducing, in an adapted reactor and so as to form a reaction medium, predetermined amounts of an organic solvent, a sugar or a sugar derivative, a compound donor of acyl groups and an enzymatic catalyst, the amount of at least one constituent of the reaction mixture being deficient, in controlled addition during the reaction of additional amounts of the deficient constituent(s), and finally purifying the resulting sucrose esters at least by separating the fine enzymatic particles from the solvent.

Abstract translation: 用于酶合成蔗糖酯的方法包括在适应的反应器中引入预定量的有机溶剂，糖或糖衍生物，酰基的化合物供体和酶催化剂 ，反应混合物中至少一种组分的量是不足的，在反应期间加入额外量的缺陷组分，最后纯化得到的蔗糖酯，至少通过从溶剂中分离出细的酶促颗粒 。

公开(公告)号：US06245535B1

公开(公告)日：2001-06-12

申请号：US09463912

申请日：2000-02-02

Applicant: Yutaka Takano ,  Masaru Hasegawa ,  Hideo Mori ,  Katsuhiko Ando ,  Keiko Ochiai ,  Hiroaki Motoyama ,  Akio Ozaki

Inventor： Yutaka Takano ,  Masaru Hasegawa ,  Hideo Mori ,  Katsuhiko Ando ,  Keiko Ochiai ,  Hiroaki Motoyama ,  Akio Ozaki

IPC: C12P762

CPC classification number: C12R1/07 ,  C12P7/62 ,  C12P17/06

Abstract: Disclosed is a process for producing a compound represented by general formula (II-a): (wherein R1 represents a hydrogen atom, a substituted or unsubstituted alkyl group, or an alkali metal; and R2 represents a substituted or unsubstituted alkyl or aryl group) [hereinafter referred to as Compound (II-a)] or the lactone form of Compound (II-a) [hereinafter referred to as Compound (II-b)] which comprises subjecting a compound represented by general formula (I-a): (wherein R1 represents a hydrogen atom, a substituted or unsubstituted alkyl group, or an alkali metal; and R2 represents a substituted or unsubstituted alkyl or aryl group) [hereinafter referred to as Compound (I-a)] or the lactone form of Compound (I-a) [hereinafter referred to as Compound (I-b)] to the action of an enzyme source derived from a microorganism belonging to the genus Bacillus and capable of converting Compound (I-a) or Compound (I-b) into Compound (II-a) or Compound (II-b) in a reaction mixture to form Compound (II-a) or Compound (II-b) in the reaction mixture, and recovering Compound (II-a) or Compound (II-b) from the reaction mixture.

Abstract translation: 公开了一种制备由通式（II-a）表示的化合物的方法：其中R 1表示氢原子，取代或未取代的烷基或碱金属; R 2表示取代或未取代的烷基或芳基） [化学式（II-a）]或化合物（II-a）的内酯形式[以下称为化合物（II-b）]] [其中包括使通式（Ia）表示的化合物 R1表示氢原子，取代或未取代的烷基或碱金属; R2表示取代或未取代的烷基或芳基）[以下称为化合物（Ia）]或化合物（Ia）的内酯形式[ （Ia）或化合物（Ib）转化为化合物（II-a）或化合物（II-a）的酶源的作用，以下称为化合物（Ib） b）在反应混合物中形成化合物（II -a）或化合物（II-b），并从反应混合物中回收化合物（II-a）或化合物（II-b）。

公开(公告)号：US06210936B1

公开(公告)日：2001-04-03

申请号：US09165043

申请日：1998-10-01

Applicant: Joseph A. Akkara ,  Ferdinando F. Bruno

Inventor： Joseph A. Akkara ,  Ferdinando F. Bruno

IPC: C12P762

CPC classification number: C12P7/625

Abstract: Protease enzyme from Bacillus subtilis and Bacillus sp. Catalyzes the acylation of organic solvent-insoluble macromolecules in isooctane solution containing vinyl esters of fatty acids, lactones or lactides as acyl donors. The reaction occurs only when the enzyme is solubilized via ion-pairing with the anionic surfactant dioctylsulfosuccinate, sodium salt (AOT). Enzyme based acylation was demonstrated in macromolecules such as silk proteins. These macromolecules are reactive either as cryogenically milled powder suspended in the organic solvent or as a thin film deposited onto ZnSe slides. This selective acylation approach represents the first attempt at using enzymes to modify organic-insoluble macromolecules in nonaqueous media.

Abstract translation: 来自枯草芽孢杆菌和芽孢杆菌的蛋白酶 催化有机溶剂不溶性大分子在含有脂肪酸，内酯或丙交酯的乙烯基酯作为酰基供体的异辛烷溶液中的酰化反应。 仅当酶通过与阴离子表面活性剂二辛基磺基琥珀酸盐钠盐（AOT）的离子配对溶解时才发生反应。 在大分子如丝蛋白中证明了酶的酰化作用。 这些大分子作为悬浮在有机溶剂中的低温研磨粉末或沉积在ZnSe载片上的薄膜是反应性的。 这种选择性酰化方法代表了在非水介质中使用酶来修饰有机不溶性大分子的第一次尝试。

公开(公告)号：US06596520B1

公开(公告)日：2003-07-22

申请号：US09610943

申请日：2000-07-06

Applicant: Thomas Friedrich ,  Rainer Stürmer

Inventor： Thomas Friedrich ,  Rainer Stürmer

IPC: C12P762

CPC classification number: C12N9/20 ,  C12N11/08 ,  Y10S435/874 ,  Y10S435/875

Abstract: Immobilized lipase is prepared by adsorbing lipase from a crude lipase solution onto polyolefin particles such as polypropylene particles which are nonpolar. The crude solution may be a cell-free culture broth. Lipase sources include Pseudomonas burkholderia and Pseudomonas aeruginosa. Uses of the immobilized lipase include enantioselective conversion of substrates such as enantioselective acylating or hydrolyzing.

Abstract translation: 通过将脂肪酶从粗脂肪酶溶液吸附到聚烯烃颗粒如非极性的聚丙烯颗粒上来制备固定的脂肪酶。 粗溶液可以是无细胞培养液。 脂肪酶来源包括铜绿假单胞菌（Pseudomonas burkholderia）和铜绿假单胞菌（Pseudomonas aeruginosa）。 固定化脂肪酶的用途包括底物的对映选择性转化，如对映选择性酰化或水解。

公开(公告)号：US06576450B2

公开(公告)日：2003-06-10

申请号：US09944243

申请日：2001-08-30

Applicant: Frank A. Skraly ,  Oliver P. Peoples

Inventor： Frank A. Skraly ,  Oliver P. Peoples

IPC: C12P762

CPC classification number: C08G8/04 ,  C12N15/52 ,  C12P7/42 ,  C12P7/625

Abstract: Organisms are provided which express enzymes such as glycerol dehydratase, diol dehydratase, acyl-CoA transferase, acyl-CoA synthetase &bgr;-ketothiolase, acetoacetyl-CoA reductase, PHA synthase, glycerol-3-phosphate dehydrogenase and glycerol-3-phosphatase, which are useful for the production of PHAs. In some cases one or more of these genes are native to the host organism and the remainder are provided from transgenes. These organisms produce poly (3-hydroxyalkanoate) homopolymers or co-polymers incorporating 3-hydroxypropionate or 3-hydroxyvalerate monomers wherein the 3-hydroxypropionate and 3-hydroxyvalreate units are derived from the enzyme catalysed conversion of diols. Suitable diols that can be used include 1,2-propanediol, 1,3 propanediol and glycerol. Biochemical pathways for obtaining the glycerol from normal cellular metabolites are also described. The PHA polymers are readily recovered and industrially useful as polymers or as starting materials for a range of chemical intermediates including 1,3-propanediol, 3-hydroxypropionaldehyde, acrylics, malonic acid, esters and amines.

Abstract translation: 提供了表达酶的生物体，例如甘油脱水酶，二醇脱水酶，酰基辅酶A转移酶，酰基辅酶A合成酶β-酮硫解酶，乙酰乙酰辅酶A还原酶，PHA合成酶，甘油-3-磷酸脱氢酶和甘油-3-磷酸酶， 对PHA的生产有用。 在一些情况下，这些基因中的一种或多种对于宿主生物体是天然的，其余的由转基因提供。 这些生物体产生聚（3-羟基链烷酸酯）均聚物或掺有3-羟基丙酸酯或3-羟基戊酸酯单体的共聚物，其中3-羟基丙酸酯和3-羟基戊酸酯单元衍生自二醇的酶催化转化。 可以使用的合适的二醇包括1,2-丙二醇，1,3-丙二醇和甘油。 还描述了从正常细胞代谢物获得甘油的生化途径。 PHA聚合物容易回收，在工业上可用作聚合物或用作一系列化学中间体（包括1,3-丙二醇，3-羟基丙醛，丙烯酸，丙二酸，酯和胺）的原料。

公开(公告)号：US06551806B1

公开(公告)日：2003-04-22

申请号：US09720914

申请日：2001-01-02

Applicant: Rainer Stürmer

Inventor： Rainer Stürmer

IPC: C12P762

CPC classification number: C12N11/08

Abstract: Enzyme-containing polymers are prepared by an anhydrous process of direct reaction of an enzyme in organic solution with crosslinking organic compounds having terminal reactive groups. In a first step, the enzyme is reacted in an organic solvent with a bifunctional monomer such as p-phenylene diisocyanate, 4-methyl-m-phenylene diisocyanate or 4,4′-methylenebisphenyl diisocyanate, and in a second step a bifunctional amine such as N-phenylethylenediamine, 1,6-diaminohexane, N,N′-diethyl-ethylenediamine or 1,4-diaminobutane is added. The enzyme-containing polymers are used as catalysts in chemical reactions such as acylation or enantioselective acylation of alcohols.

Abstract translation: 含酶聚合物通过无机方法通过酶在有机溶液中与具有末端反应性基团的交联有机化合物直接反应来制备。 在第一步中，酶在有机溶剂中与双官能单体如对亚苯基二异氰酸酯，4-甲基间苯二异氰酸酯或4,4'-亚甲基二苯基二异氰酸酯反应，在第二步中，将双官能胺如 作为N-苯乙二胺，1,6-二氨基己烷，N，N'-二乙基 - 乙二胺或1,4-二氨基丁烷。 含酶聚合物用作化学反应中的催化剂，例如醇的酰化或对映选择性酰化。